A novel attachment mechanism for Burkholderia cepacia complex

项目总结 细菌对宿主细胞的附着是致病过程中的关键步骤。因此，对细菌的预防 附着于宿主细胞是阻止疾病进展的理想靶点。严重缺乏对这一问题的了解 伯克霍尔德氏菌的附着机制极大地抑制了我们开发有效治疗的能力 战略。洋葱伯克霍尔德氏菌复合体(BCC)，由20多种革兰氏阴性菌组成， 对公众健康构成重大威胁。基底细胞癌在囊性纤维化(CF)中引起严重的机会性肺部感染 世界各地的患者。CF是一种危及生命的疾病，无法治愈和治疗与CF相关的感染 很难。美国大约有3万名CF患者，全球有超过7万人。 基底细胞癌也是一组重要的病原菌，导致医院感染中的其他免疫抑制 以几次感染暴发为突出表现的个人。 伪颌伯克霍尔德氏菌一种新型表面附着蛋白的重大研究发现 (Sap1Bp)是一种以前未被描述的假设蛋白质，它为研究一种新的 而且，看起来，许多伯克霍尔德氏菌物种利用了普遍的依附机制。Sap1Bp在功能上 在许多伯克霍尔德氏菌物种中保守，包括来自BCC临床相关成员的Sap1(Sap1Bcc)。 BCC使用的依附机制很多，而且很复杂。但是，没有关联的 到目前为止，来自宿主细胞的受体伙伴已经被发现，在 了解BCC中的依附机制。我们发现Sap1Bcc对BCC有显著贡献 附着在宿主细胞上，是多种动物模型中完整的基底细胞癌发病所必需的。此外， 我们发现Sap1宿主细胞受体是糖蛋白非转移性黑色素瘤蛋白B(GPNMB)， 它与多种癌症和阿尔茨海默病有关，揭示了一种新颖而独特的相互作用 从未被任何细菌病原体利用来附着和入侵宿主细胞。 在这一应用中，我们建议探索这种重要的表面附着蛋白的机制。 Sap1Bcc与其宿主细胞受体GPNMB的体外相互作用及其在体内的重要性 发病机制。目标1A将在分子水平上表征Sap1Bcc和GPNMB的相互作用，通过确定 它们的相互作用亲和力和确定依恋功能的关键残基，这对未来的研究至关重要。 它的致病机制。目的1B试图从功能上表征GPNMB在Sap1Bcc依赖中的作用 Bcc附着于GPNMB基因敲除和过表达细胞。AIM 1C将调查GPNMB在 BCC小鼠感染后新毒力因子Sap1Bcc与宿主细胞的相互作用 受体GPNMB直接参与疾病的发生。新发现的Sap1Bcc的详细特征 /GPNMB相互作用可能提供潜在的药物靶点和替代治疗策略 基底细胞癌感染的治疗结果。

PROJECT SUMMARY Bacterial attachment to host cells is a critical step for pathogenesis. Thus, the prevention of bacterial attachment to host cells is an ideal target to halt disease progression. A severe lack of knowledge of the attachment mechanism of Burkholderia species greatly inhibits our ability to develop effective treatment strategies. Burkholderia cepacia complex (Bcc), consisting of more than 20 species of Gram-negative bacteria, poses a major threat to public health. Bcc causes severe opportunistic lung infections in cystic fibrosis (CF) patients worldwide. CF is a life-threatening condition with no cure and treatment of CF-related infections is difficult. There are about 30,000 people with CF in the United States and more than 70,000 people worldwide. Bcc is also a group of significant pathogens leading to nosocomial infections in other immunosuppressed individuals highlighted by several outbreaks of infection. Our significant research discovery of a novel surface attachment protein from Burkholderia pseudomallei (Sap1Bp), a previously uncharacterized hypothetical protein, has provided an opportunity to investigate a new and, seemingly, universal attachment mechanism exploited by many Burkholderia species. Sap1Bp is functionally conserved across many Burkholderia species, including Sap1 from clinically relevant members of Bcc (Sap1Bcc). There are many attachment mechanisms employed by Bcc and they are complex. However, no associated receptor partners from host cells have been discovered to date, leaving significant knowledge gaps in the understanding of attachment mechanisms in the Bcc. We found that Sap1Bcc contributes significantly to Bcc attachment to the host cells and is required for complete Bcc pathogenesis in multiple animal models. Moreover, we discovered the Sap1 host-cell receptor to be the glycoprotein non-metastatic melanoma protein B (GPNMB), which is associated with various cancers and Alzheimer's disease, revealing a novel and unique interaction that has never been exploited by any bacterial pathogens to attach and invade host cells. In this application, we proposed to explore the mechanisms of this important surface attachment protein Sap1Bcc to interact with its host-cell receptor GPNMB in vitro and the importance of this interaction for in vivo pathogenesis. Aim 1A will characterize Sap1Bcc and GPNMB interaction at the molecular level by determining their interaction affinity and defining the key residues for attachment function, which is critical for future study of its virulence mechanism. Aim 1B seeks to functionally characterize the roles of GPNMB in Sap1Bcc-dependent Bcc attachment to GPNMB knockout and overexpression cells. Aim 1C will investigate the role of GPNMB during Bcc murine infection to determine if the interaction between the novel virulence factor Sap1Bcc and its host cell receptor GPNMB directly contributes to disease. Detailed characterizations of this newly discovered Sap1Bcc /GPNMB interaction could provide potential drug-target and alternative therapeutic strategies toward better treatment outcomes of Bcc infection.